Age-dependent impairment of angiogenesis leads to increased progression of ischemic vascular diseases, delayed wound healing, as well as slower tumor growth and spread. The mechanisms of reduced angiogenesis have not been fully elucidated yet but may include alterations in endothelial responsiveness, cell-matrix interactions and lack of angiogenic factors and their receptors. Neuropeptide Y (NPY) is commonly known as a sympathetic transmitter and vasoconstrictor. Recent our laboratory has established its new and potent activities as an angiogenic and mitogenic factor for vascular smooth muscle and endothelial cells in vitro, and in vivo in the rat hindlimb ischemic model. The overall goal of this research is to establish if alternations in the NPY system are involved in the age-related impairment of angiogenesis. The specific aims are to determine if: 1) NPY-mediated angiogenesis is diminished with age; 2) NPY synthesis/release occurring under physiological condition and in response to angiogenic stimuli is reduced in aged animals; 3) expression of the NPY angiogenic receptors (Y1, Y2, Y5) and its converting enzyme dipeptidyl peptidase IV (DPPIV, which converts NPY to an angiogenically active fragment, NPY3-36) is altered with age 4) age-related changes in NPY angiogenic systems are due to decreased levels of basic fibroblast growth factor (bFGF) in aging. These studies will be perform4ed on both animal (young and aged mice) and cellular models. Spontaneous, as well as NPY- and bFGF-induced sprouting of mouse aortas will be compared between both age groups. The mouse hindlimb ischemic model will be employed to determine the effect of aging on NPY-induced angiogenesis in ischemia. New vessels will be detected by CD31 immunostaining and by their casting by perfusion with latex. Systemic and local NPY levels, as well as expression of angiogenic stimuli, will be determined. The cellular model will include NPY stimulation of aortic endothelial cells from young and aged mice and measuring their mitogenic response in parallel to the expression of NPY receptors. Interactions between bFGF and NPY angiogenic systems and their age-related modulations will be established by determining the levels of growth factors and NPY receptors in ischemic tissues of aged and young animals, also after NPY or bFGF treatment. Moreover, cross-inhibition with neutralizing antibody of bFGF- and NPY-induced neovascularization as well as bFGF-mediated angiogenesis in Y2 knockout mice will be investigated using aortic sprouting assay. This research will help to establish the mechanism(s) of age-related changes in angiogenesis, which may have important implications for the development of ne2w therapeutic strategies in older patients-an enhancement of angiogenesis in the ischemic disease.